P879: EXENT AND FLC MASS SPECTROMETRY FOR SEROLOGICAL IDENTIFICATION AND QUANTIFICATION OF MONOCLONAL IMMUNOGLOBULINS IN MULTIPLE MYELOMA
Authors: S. Huhn, A. M. Poos, E. K. Mai, M. Hänel, H. J. Salwender, J. Dürig, I. W. Blau, C. Scheid, K. C. Weisel, M. Munder, O. Berlanga, U. Bertsch, H. Goldschmidt, N. Weinhold
Journal: HemaSphere (2022)

Background: Mass spectrometry has broad potential for screening and monitoring monoclonal gammopathies. The technology overcomes some of the limitations inherent to standard electrophoretic methods for identifying monoclonal immunoglobulins and is amenable to automation. Aims: Here we assess the performance of the EXENT assays for the quantification of M-proteins and whether free light chain evaluation by mass spectrometry (FLC-MS) offers added sensitivity during multiple myeloma monitoring. Methods: The MM5 trial compared two regimens of bortezomib-based induction therapy and of lenalidomide consolidation followed by lenalidomide maintenance in newly diagnosed multiple myeloma patients. We used mass spectrometry for identifying serum M-proteins in samples at baseline (n=434), post-induction (n=340), and post-consolidation (n=294). Briefly, polyclonal antibodies (anti-IgG,-IgA,-IgM,-total κ and -total λ [EXENT, The Binding Site, UK] and anti-free κ and -free λ [FLC-MS]) covalently attached to paramagnetic microparticles were separately incubated with serum, washed, treated to elute and reduce patient immunoglobulins, and spotted onto MALDI plates with HCCA matrix. Spectra were generated using the MALDI-TOF-MS system. Light chain mass spectra were obtained using EXENT software to yield immunoglobulin isotype, mass-to-charge ratio (m/z) and quantity (g/L). Results were compared to serum protein electrophoresis, immunofixation, and FLC ratios (Freelite®, The Binding Site, UK). Results: EXENT demonstrated improved sensitivity over immunofixation for identifying the M-protein at each time-point of the MM5 trial: 99% vs. 95% at baseline; 91% vs 80% post-induction; and 63% vs. 46% post-consolidation. Agreement between methods was 87%. The addition of FLC-MS to EXENT further improved sensitivity to 100%, 95.0%, and 72% at each respective time-point, whereas the combination of immunofixation and FLC ratios resulted in sensitivities of 100%, 90%, and 54%. Overall concordance between EXENT+FLC-MS and the combination of immunofixation and FLC ratios was 100% at baseline, 93% post-induction, and 73% post-consolidation. In all, 158 (11%) samples had an M-protein by EXENT+FLC-MS only, and 28 (2%) by the combination of immunofixation and FLC ratios. 18 and 92 patients achieved an IMWG complete response after induction and consolidation, respectively. EXENT identified an M-protein in 5 (28%), and 23 (25%) of these patients, whereas EXENT+FLC-MS was more sensitive and detected monoclonal proteins in 11 (61%), and 38 (41%) patients, respectively. There was acceptable quantitative agreement for M-protein measurements between EXENT and serum protein electrophoresis (n=631: Passing-Bablok fit: y=0.85x-0.58; coefficient of determination R2= 0.89; Bland-Altman relative bias (95%CI): -32.62% (-123.79-58.55%)). However, there was a substantial discordance in 285 samples <10g/L by serum protein electrophoresis (y=0.58x+0.54; R2=0.28; -44.84% (-179.29 – 84.60%)) compared to 346 samples >10g/L (y=0.85x-0.25; R2=0.82; -19.91% (95%CI: -97.32 – 23.56%)); suggesting overestimation by serum protein electrophoresis at lower levels. Summary/Conclusion: EXENT provides superior performance for the identification of M-proteins throughout monitoring. Sensitivity can be improved by incorporating FLC-MS assessments, thus providing a means for determining serological residual disease in patients at complete response by IMWG criteria. Quantification of the M-protein by mass spectrometry compares well to standard methods and offers accurate results at levels below the analytical threshold of confidence for electrophoretic approaches.